Several antioxidant small molecules that are synthesized by plants are essential vitamins in the human diet. These nutrients, including vitamin A precursors (carotenoids), vitamin E (tocopherols), and vitamin C (ascorbate), have critical functions in quenching and scavenging reactive oxygen species that are inevitable byproducts of metabolism in all aerobic organisms, from bacteria to plants and animals. Despite the importance of antioxidant vitamins for human health, many fundamental questions remain unanswered about their function and the regulation of their biosynthesis. This proposal addresses several of these questions using molecular genetic, genomic, and physiological approaches in a tractable, unicellular green alga, Chlamydomonas reinhardti. Chlamydomonas represents a powerful eukaryotic model organism for studying antioxidant metabolism in plants, much like the budding yeast Saccharomyces cerevisiae is used as a model for animals. The specific aims of this proposal are (1) to complete a molecular and genetic analysis of carotenoid biosynthesis in Chlamydomonas, (2) to generate a series of mutants affecting tocopherol composition in order to test the antioxidant function of different tocopherols in vivo, (3) to examine functional overlap between carotenoids and tocopherols as antioxidants in polyunsaturated lipid membranes, and (4) to perform genomic and genetic analyses of how carotenoid and tocopherol levels are regulated in cells by environmental factors, such as light intensity. This investigation will provide new insights into the roles of lipid-soluble antioxidants in preventing age-related diseases such as macular degeneration in humans and will provide a foundation for rational modification of carotenoid and tocopherol composition and content in plants to benefit human nutrition and health.